Project Summary/Abstract This proposal addresses how cells communicate with each other to control development by elucidating the signaling components, modulators and mechanisms that govern cell-cell interactions. Understanding the underlying genetic circuitry and molecular mechanisms is critical, because aberrant activity of these same signaling pathways have profound effects on human health, most notably as causal agents of cancer, congenital defects and diverse physiological disorders. Regulating signaling appropriately?in space or cell population, in time, strength or duration, or in combination with other signaling inputs?is thus crucial both for normal development and for insight into human disease. This proposal builds on foundational work on two key developmental paradigms in C. elegans: (i) specification of alternative cell fates by LIN- 12/Notch signaling in the gonad and (ii) the integration of LIN-12/Notch and EGFR-Ras-ERK signaling to pattern three distinct fates in the vulva. Although each paradigm has unique attributes, they provide a unified platform for elucidating regulatory circuitry and mechanisms underlying cell fate decisions and the function and regulation of major, conserved signaling systems. These C. elegans paradigms have a distinguished record of fostering discoveries that are directly applicable to basic human developmental biology and medicine, and a major reason is that they are especially amenable to genetic analysis. The proposed work will utilize a combination of classical genetic approaches and CRISPR/Cas9-engineering and other technologies to manipulate and monitor gene function, as well new microfluidic-based methods for longitudinal imaging and automated data collection that allows unprecedented cellular precision and temporal resolution in analyzing cell fate choice, gene expression, and signaling dynamics. The work will address three key gaps in understanding: (i) the relationship between signaling dynamics and cell fate specification, (ii) the regulatory circuitry and mechanisms that enable precise and robust spatial patterning, and (iii) the regulation of developmental progression through integrating life history and environmental cues with spatial patterning. The identification of mechanisms by which major, conserved signaling systems are regulated and integrated with each other is increasingly important in the era of personalized medicine and the deeper understanding of developmental mechanism we will achieve through these studies will be potentially applicable to developing diagnostic and therapeutic tools for human disease.